275 .
TREPONEMA SUCCINIFACIENS MEDIUM
What is this page?
Equipment needed:
Autoclave
Centrifuge
Gassing station
Hungate tubes
Final pH:
7.4
Select a strain (optional):
ml
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
Solution A | 920 | ml | ||
|
|
Solution B | 50 | ml | ||
|
|
Solution C | 30 | ml | ||
| 1 Dissolve ingredients of solution A and sparge with 80% N2 and 20% CO2 gas atmosphere to make it anoxic. Then distribute medium under the same gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. Solutions B is prepared under 100% N2 gas atmosphere and autoclaved. Solution C is prepared under 80% N2 and 20% CO2 gas atmosphere and autoclaved. To complete the medium appropriate amounts of solutions B and C are added to the sterile solution A in the sequence as indicated. Final pH of the medium should be at 7.4. | |||||
Solution A #
920
| Compound | Amount | Unit | |
|---|---|---|---|
|
|
CaCl2 x 2 H2O | 0.1 | g |
|
|
MgSO4 x 7 H2O | 0.1 | g |
|
|
KH2PO4 | 0.5 | g |
|
|
K2HPO4 | 0.5 | g |
|
|
NaCl | 1.0 | g |
|
|
Clarified rumen fluid | 300.0 | ml |
|
|
Yeast extract
(OXOID) |
0.5 | g |
|
|
Peptone
(BD BACTO) |
0.5 | g |
|
|
(NH4)2SO4 | 0.5 | g |
|
|
Sodium resazurin
(0.1% w/v) |
0.5 | ml |
|
|
L-Cysteine HCl x H2O | 0.5 | g |
|
|
Distilled water | 620.0 | ml |
Clarified rumen fluid #
1000
| 1 Rumen fluid from cow or sheep (obtained from fistulated animals or abattoir refuse) is filtered through muslin, autoclaved at 121°C for 15 min and then centrifuged at 27,000 g for 20 min. The supernatant is made anoxic by sparging with 100% N2 gas for 15 min, dispensed under same gas atmosphere into anoxic serum vials to 30% of volume and then stored frozen at -20°C. |